Epigenetic restoration and activation of ERβ: an inspiring approach for treatment of triple-negative breast cancer

Abstract

Background: Triple-negative breast cancer (TNBC) is one of the most aggressive subtypes of breast cancer. TNBC lacks targeted therapy receptors, rendering endocrine and HER2-targeted therapies ineffective. TNBC is typically treated with cytotoxic chemotherapy followed by surgery. Targeting epigenetic modifications could potentially be a new effective TNBC target therapy. The aim of this study is to examine the effects of epigenetic drugs, decitabine as DNA methyltransferase inhibitor (DNMTI) and vorinostat as histone deacetylase inhibitor (HDACI), and the ERβ agonist DPN on ERα and ERβ re-expressions in the MDA-MB-231 cells as a model of TNBC.

Methods: Using MTT assay, the IC₅₀ of decitabine, vorinostat, and DPN on MDA-MB-231 cells were determined. The effects of all drugs alone or in combinations on MDA-MB-231 cells were evaluated. qRT-PCR was used to determine ERα & ERβ gene expression. Caspase-3 activity and the protein expression levels of VEGF, Cyclin D1, and IGF-1 were assessed.

Results: Both ERα and ERβ mRNA were re-expressed in different high levels in all treated groups, especially in the triple therapy group compared with control. Significantly, the triple drugs therapy showed the lowest levels of VEGF, Cyclin D1, and IGF-1 and the highest level of Caspase-3 activity, indicating a possible antitumor effect of ERβ activation through decreasing proliferation and angiogenesis and increasing apoptosis in MDA-MB-231 cells.

Conclusions: The antiproliferative effect of ERβ could be retained when co-expressed with Erα using a powerful epigenetic combination of Decitabine and vorinostat with DPN.

Keywords: DNMTI; DPN; Decitabine; ERβ agonist; HDACI; MDA-MB-231; TNBC; Vorinostat.

Fig. 1

Sigmoidal curve for MTT assay showing IC₅₀ values and the inhibition % of decitabine (A), vorinostat (B), and DPN (C) on MDA-MB-231 cells. Each data point represents an average of three independent experiments

Fig. 2

Fold changes of ERα (A) and ERβ (B) relative expression levels relative to the control group after treatment with decitabine (4 µM), vorinostat (0.26 µM), DPN (0.093 µM) and their combinations for 72 h in MDA-MB-231 cells

Fig. 3

Effects of decitabine (4 µM), vorinostat (0.26 µM), DPN (0.093 µM) and their combinations treatments on the treated groups of MDA-MB-231 cells A cyclin D1, B IGF-1 C active caspase-3 activity, and D VEGF biomarkers. Data are presented as the mean ± SEM of three samples each performed in triplicate. Statistically significant differences between groups (p < 0.05) are designated as *significant vs. control, ^πsignificant vs. DPN, ^#significant vs. decitabine, ^$significant vs. vorinostat, ^Δsignificant vs. decitabine & vorinostat